Within the pharmaceutical industry, there is a significant number of compounds, available in compound repositories, through the use of combinatorial chemistry. Hits from these compound libraries are identified through the use of high throughput screening (HTS). The primary function of HTS is to test various chemical compounds from a compound repository against multiple disease targets in many different biological assays. Traditional HTS commonly utilizes 96-well microtiter plates. There is a large incentive in the pharmaceutical industry to miniaturize these microplate assays to reduce cost, reduce waste, and speed up timelines. There has been a change from the 96-well format to higher well densities such as 384- and 1536-well formats, however these can present challenges in relation to liquid handling, signal detection instrumentation, and assay technology. In addition, typical difficulties encountered when using microtiter plate based screening assays include e.g. (i) differential growth and/or gene expression of identical clones in separate wells of the microtiter plate and (ii) differential stress exposure (e.g. heat treatment, humidity) across the plate.
Efforts have been made towards resolving the aforementioned difficulties. For example, WO 99/35496 provides a method and apparatus for high density format screening for bioactive molecules with a much simplified technique for test compound delivery to a layer of cells, i.e. without the need of complicated fluid handling. In the said method, up to 6144 test compounds may be simultaneously screened for bioactivity.
Nevertheless, there is a need for even higher density cell-based functional biological assays which remain one of the most difficult types of assays to miniaturize due to the limitations of delivering microliter quantities of cells consistently without shearing the cells or activating stress responses of the cells themselves, therefore interfering with the biological assay.
With the advent of combinatorial chemistry approaches to identify pharmacologically useful compounds, it is increasingly evident that there is a need for methods and apparatuses at microarray levels, capable of performing high throughput characterization of pharmacological profiles and corresponding potencies of the compounds in synthesized combinatorial libraries.
Microarrays of living cells could provide a shortcut to the development of safer and more customized personal drugs and a better understanding of the molecular pathways encountered in the functioning of cellular organisms. As an example, the Whitehead Institute for Biomedical Research in Cambridge developed microarrays of living cells for high-throughput analysis of gene function in mammalian cells (Nature, Vol. 411, 3 May 2001). While said technology is highly effective, there are, however, a number of limitations to its use including, the relative high reagent quantities needed of which a substantial amount is never in contact with the array and is therefore wasted, and the requirement of cumbersome and time-consuming handlings.
U.S. Pat. No. 6,103,479 discloses a further example of a microarray of living cells. The disclosed miniaturized cell arrays characterized by a reduced well and array size allow for high content screening. The non-porous character, however, does not allow the cells growing in/on the wells or cell binding sites on the array to overcome spreading under standard growth conditions which obviously interferes with proper cell sample discrimination.
As will be appreciated in the art, there is a continuous need for improved methods which overcome the aforementioned disadvantages.
It is therefore an object of the present invention to provide a highly efficient and cost-effective method for integrated cell-based assays using microarrays.
It is a further object of the present invention to provide a method for high-throughput cell-based assays requiring minimal amounts of sample and reagents.
It is also an object of the present invention to provide microarrays or kits to perform such methods.